The enediyne antibiotics are currently the focus of intense research activity in the fields of chemistry, biology, and medical sciences, because of their unique molecular architecture, biological activities, and modes of actions (Doyle and Borders (1995) Enediyne antibiotics as antitumor agents. Marcel-Dekker, New York, Thorson et al. (1999) Bioorg. Chem., 27: 172–188). Since the unveiling of the structure of neocarzinostatin chromophore (Edo et al. (1985) Tetrahedron Lett. 26: 331–340) in 1985, the enediyne family has grown steadily. Thus far, there have been three basic groups within the enediyne antibiotic family: (a) the calicheamicin/esperamicin type, which includes the calicheamicins, the esperamicins, and namenamicin, (b) the dynemicin type, and (c) the chromoprotein type, consisting of an apoprotein and an unstable enediyne chromophore. The latter group includes neocarzinostatin, kedarcidin, C-1027 (FIG. 1), and maduropeptin, whose enediyne chromophore structures have been established, as well as several others whose enediyne chromophore structures are yet to be determined due to their instability (Thorson et al. (1999) Bioorg. Chem., 27: 172–188). N1999A2, in contrast to the other chromoproteins, exists as an enediyne chromophore alone despite the fact that its structure is very similar to the other chromoprotein chromophore (Ando et al.(1998) Tetra. Letts., 39: 6495–6480).
As a family, the enediyne antibiotics are the most potent, highly active antitumor agents ever discovered. Some members are 1000 times more potent than adriamycin, one of the most effective, clinically used antitumor antibiotics (Zhen et al. (1989) J. Antibiot. 42: 1294–1298). All members of this family contain a unit consisting of two acetylenic groups conjugated to a double bond or incipient double bond within a nine or ten-membered ring; i.e., the enediyne core as exemplified by C-1027 in FIG. 1. As the consequence of this structural feature, these compounds share a common mechanism of action: the enediyne core undergoes an electronic rearrangement to form a transient benzenoid diradical, which is positioned in the minor groove of DNA so as to damage DNA by abstracting hydrogen atoms from deoxyriboses on both strands (FIG. 1). Reaction of the resulting deoxyribose carbon-centered radicals with molecular oxygen initiates a process that results in both single-strand and double-strand DNA cleavages (Doyle and Borders (1995) Enediyne antibiotics as antitumor agents. Marcel-Dekker, New York; Ikemoton et al. (1995) Proc. Natl. Acad. Sci. USA 92:10506–10510; Myers et al. (I 997) J. Am. Chem. Soc. 119: 2965–2972; Stassinopoulos et al. (1996) Science 272: 1943–1946; Thorson et al. (1999) Bioorg. Chem., 27: 172–188; Xu et al. (1997) J. Am. Chem. Soc. 119: 1133–1134). This novel mechanism of DNA damage has important implications for their application as potent cancer chemotherapeutic agents (Doyle and Borders (1995) supra.; Sievers et al. (1999) Blood 93: 3678–3684).
As an alternative to making structural analogs of microbial metabolites by chemical synthesis, manipulations of genes governing secondary metabolism offer a promising alternative allowing preparation of these compounds biosynthetically (Cane et al. (1998) Science 282: 63–68; Hutchinson and Fujii. (1995) Ann. Rev. Microbiol. 49: 201–38; Katz and Donadio (1993) Ann. Rev. Microbiol. 47: 875–912). The success of the latter approach depends critically on the availability of novel genetic systems and on genes encoding novel enzyme activities. The enediynes offer a distinct opportunity to study the biosynthesis of their unique molecular scaffolds and the mechanism of self-resistance to extremely cytotoxic natural products. Elucidation of these aspects provides access to rational engineering of enediyne biosynthesis for novel drug leads and makes it possible to construct enediyne overproducing strains by de-regulating the biosynthetic machinery. In addition, elucidation of an enediyne gene cluster contributes to the general field of combinatorial biosynthesis by expanding the repertoire of novel polyketide synthase (PKS) and deoxysugar biosynthesis genes as well as other genes uniquely associated with enediyne biosynthesis, leading to the making of novel enediynes via combinatorial biosynthesis.